The long-term goals of this work are to understand the mechanisms that generate and regulate motility in cilia and flagella. Human mutations that inhibit motility of these organelles cause upper respiratory illnesses and infertility, and related forms of motility are important for mitosis and directed transport of many intracellular particles. Flagellar dynein ATPases are large, multi-subunit motor enzymes that power cilia and flagella. Most flagella have two non-identical rows of dyneins attached to each doublet microtubule, both of which contribute to microtubule sliding and flagellar motility. Dyneins of the outer row more closely resemble cytoplasmic dyneins in containing two or three ca. 500 kD catalytic heavy chains, two ca. 75 kD intermediate chains and multiple light chains. While the role of dyneins as microtubule-associated motors for both ciliary and cytoplasmic motility is well recognized, little is known about their regulation or their mode of attachment to the loads they carry (vesicles or chromosomes in the cytoplasm, doublet microtubules in cilia and flagella). Goals of the proposed experiments are to characterize interactions between dynein subunits and doublet microtubule-associated proteins that are important for dynein attachment during flagellar assembly and for regulation of outer row dynein motors. Using the outer row dynein of Chlamydomonas reinhardtii flagella as a model system, genes essential for normal dynein assembly and function are being cloned, sequenced, and genetically mapped. The function of each gene product in cytoplasmic assembly, targeting to flagella, and in situ regulation is examined using molecular cloning, electron microscopic and biochemical approaches. Experiments proposed here focus on the products of four genes identified through published and preliminary studies of assembly mutations as having unique roles in dynein assembly: oda7, oda8, oda15 and pfl3. All materials are now available for rapid progress in determining the products of these genes, which should provide key information about mechanisms that regulate the intracellular distribution and activity of flagellar dynein motors.